Association of serum adiponectin and leptin levels with renal function in kidney transplant recipients with or without new-onset diabetes after transplantation.

PURPOSE: To evaluate serum adiponectin and leptin concentration in new-onset diabetes after transplantation (NODAT) and non-NODAT patients and association with renal function in kidney transplant recipients (KTRs). PATIENTS AND METHODS: A study of 314 consecutive adults KTRs divided into four groups: 236 individuals without NODAT who had renal insufficiency (RI; n = 56) or normal renal function (n = 180) and 78 patients with NODAT who had RI (n = 17) or normal renal function (n = 61). NODAT was diagnosed based on venous fasting blood glucose or HbA1c with the criteria of the American Diabetes Association. Renal insufficiency was defined according to KDOQI 2002 guidelines. RESULTS: In the NODAT group, the median level of serum adiponectin was lower than that of non-NODAT one (30 µg/ml vs 37.15 µg/ml, p < 0.001); in contrast, the median leptin concentration was higher (4.27 ng/ml vs 4.05 ng/ml, p = 0.024). In the RI group, both median serum adiponectin and leptin levels were higher than those of non-RI one (Adiponectin: 40.01 µg/ml vs 33.7 µg/ml; Leptin: 4.51 ng/ml vs 3.91 ng/ml, p < 0.001 both). We found that BMI was related to both adiponectin and leptin levels in both NODAT, non-NODAT, and all subject groups, based on univariate and multivariate linear regression analysis. CONCLUSION: New-onset diabetes after transplantation, BMI, and renal insufficiency were affected to the serum level of adiponectin and leptin in KTRs.

Serum hs-CRP measured prior transplantation predicts of new-onset diabetes after transplantation in renal transplant recipients.

BACKGROUND: To assess the incidence of new-onset diabetes after transplantation (NODAT) for the first year post-transplantation and the predictive value of high-sensitivity C-reactive Protein (hs-CRP) before transplantation for NODAT prediction in kidney transplantation patients. MATERIAL AND METHODS: A study of 251 consecutive adult end-stage kidney disease patients transplanted kidneys from living donors, follow-up during the first year to find NODAT. We diagnosed NODAT based on blood glucose or HbA1c following to the criteria of the American Diabetes Association. RESULTS: The ratio of NODAT was 12.4%. The mean age, mean BMI, the proportion of arteriosclerosis, and the median hs-CRP level in NODAT group were significantly higher than those of non-NODAT group with p < 0.05. Age, BMI and serum hs-CRP had a predictive value for NODAT (Age: AUC = 0.62, p < 0.05, BMI: AUC = 0.626, hs-CRP: AUC = 0.748, p < 0.001). CONCLUSION: Serum hs-CRP level measured prior transplantation is a good predictor for NODAT in renal transplant recipients.

A metagenome-derived thermostable ß-glucanase with an unusual module architecture which defines the new glycoside hydrolase family GH148.

The discovery of novel and robust enzymes for the breakdown of plant biomass bears tremendous potential for the development of sustainable production processes in the rapidly evolving new bioeconomy. By functional screening of a metagenomic library from a volcano soil sample a novel thermostable endo-ß-glucanase (EngU) which is unusual with regard to its module architecture and cleavage specificity was identified. Various recombinant EngU variants were characterized. Assignment of EngU to an existing glycoside hydrolase (GH) family was not possible. Two regions of EngU showed weak sequence similarity to proteins of the GH clan GH-A, and acidic residues crucial for catalytic activity of EngU were identified by mutation. Unusual, a carbohydrate-binding module (CBM4) which displayed binding affinity for ß-glucan, lichenin and carboxymethyl-cellulose was found as an insertion between these two regions. EngU hydrolyzed ß-1,4 linkages in carboxymethyl-cellulose, but displayed its highest activity with mixed linkage (ß-1,3-/ß-1,4-) glucans such as barley ß-glucan and lichenin, where in contrast to characterized lichenases cleavage occurred predominantly at the ß-1,3 linkages of C4-substituted glucose residues. EngU and numerous related enzymes with previously unknown function represent a new GH family of biomass-degrading enzymes within the GH-A clan. The name assigned to the new GH family is GH148.

Novel archaeal thermostable cellulases from an oil reservoir metagenome.

Microbial assemblages were sampled from an offshore deep sub-surface petroleum reservoir 2.5 km below the ocean floor off the coast of Norway, providing conditions of high temperature and pressure, to identify new thermostable enzymes. In this study, we used DNA sequences obtained directly from the sample metagenome and from a derived fosmid library to survey the functional diversity of this extreme habitat. The metagenomic fosmid library containing 11,520 clones was screened using function- and sequence-based methods to identify recombinant clones expressing carbohydrate-degrading enzymes. Open reading frames (ORFs) encoding carbohydrate-degrading enzymes were predicted by BLAST against the CAZy database, and many fosmid clones expressing carbohydrate-degrading activities were discovered by functional screening using Escherichia coli as a heterologous host. Each complete ORF predicted to encode a cellulase identified from sequence- or function-based screening was subcloned in an expression vector. Five subclones was found to have significant activity using a fluorescent cellulose model substrate, and three of these were observed to be highly thermostable. Based on phylogenetic analyses, the thermostable cellulases were derived from thermophilic Archaea and are distinct from known cellulases. Cellulase F1, obtained from function-based screening, contains two distinct cellulase modules, perhaps resulting from fusion of two archaeal cellulases and with a novel protein structure that may result in enhanced activity and thermostability. This enzyme was found to exhibit exocellulase function and to have a remarkably high activity compared to commercially available enzymes. Results from this study highlight the complementarity of hybrid approaches for enzyme discovery, combining sequence- and function-based screening.

Functional Screening of Hydrolytic Activities Reveals an Extremely Thermostable Cellulase from a Deep-Sea Archaeon.

Extreme habitats serve as a source of enzymes that are active under extreme conditions and are candidates for industrial applications. In this work, six large-insert mixed genomic libraries were screened for hydrolase activities in a broad temperature range (8-70°C). Among a variety of hydrolytic activities, one fosmid clone, derived from a library of pooled isolates of hyperthermophilic archaea from deep sea vents, displayed hydrolytic activity on carboxymethyl cellulose substrate plates at 70°C but not at lower temperatures. Sequence analysis of the fosmid insert revealed a gene encoding a novel glycoside hydrolase family 12 (GHF12) endo-1,4-ß-glucanase, termed Cel12E. The enzyme shares 45% sequence identity with a protein from the archaeon Thermococcus sp. AM4 and displays a unique multidomain architecture. Biochemical characterization of Cel12E revealed a remarkably thermostable protein, which appears to be of archaeal origin. The enzyme displayed maximum activity at 92°C and was active on a variety of linear 1,4-ß-glucans like carboxymethyl cellulose, ß-glucan, lichenan, and phosphoric acid swollen cellulose. The protein is able to bind to various insoluble ß-glucans. Product pattern analysis indicated that Cel12E is an endo-cleaving ß-glucanase. Cel12E expands the toolbox of hyperthermostable archaeal cellulases with biotechnological potential.

Characterization of chromosomal and megaplasmid partitioning loci in Thermus thermophilus HB27.

BACKGROUND: In low-copy-number plasmids, the partitioning loci (par) act to ensure proper plasmid segregation and copy number maintenance in the daughter cells. In many bacterial species, par gene homologues are encoded on the chromosome, but their function is much less understood. In the two-replicon, polyploid genome of the hyperthermophilic bacterium Thermus thermophilus, both the chromosome and the megaplasmid encode par gene homologues (parABc and parABm, respectively). The mode of partitioning of the two replicons and the role of the two Par systems in the replication, segregation and maintenance of the genome copies are completely unknown in this organism. RESULTS: We generated a series of chromosomal and megaplasmid par mutants and sGFP reporter strains and analyzed them with respect to DNA segregation defects, genome copy number and replication origin localization. We show that the two ParB proteins specifically bind their cognate centromere-like sequences parS, and that both ParB-parS complexes localize at the cell poles. Deletion of the chromosomal parAB genes did not apparently affect the cell growth, the frequency of cells with aberrant nucleoids, or the chromosome and megaplasmid replication. In contrast, deletion of the megaplasmid parAB operon or of the parB gene was not possible, indicating essentiality of the megaplasmid-encoded Par system. A mutant expressing lower amounts of ParABm showed growth defects, a high frequency of cells with irregular nucleoids and a loss of a large portion of the megaplasmid. The truncated megaplasmid could not be partitioned appropriately, as interlinked megaplasmid molecules (catenenes) could be detected, and the ParBm-parSm complexes in this mutant lost their polar localization. CONCLUSIONS: We show that in T. thermophilus the chromosomal par locus is not required for either the chromosomal or megaplasmid bulk DNA replication and segregation. In contrast, the megaplasmid Par system of T. thermophilus is needed for the proper replication and segregation of the megaplasmid, and is essential for its maintenance. The two Par sets in T. thermophilus appear to function in a replicon-specific manner. To our knowledge, this is the first analysis of Par systems in a polyploid bacterium.